Regulatory T cells (Tregs) are key modulators of inflammation and are important for the maintenance of peripheral tolerance. Adoptive immunotherapy with polyclonal Tregs holds promise in organ transplantation, graft-versus-host disease, and autoimmune diseases, but may be enhanced by antigen-specific, long-lived Treg cells. We modified primary human Tregs with chimeric antigen-receptors (CARs) bearing different costimulatory domains and performed in vitro analyses of their phenotype and function. While neither the presence of a CAR nor the type of costimulation domain influenced Foxp3 expression in Tregs, the costimulation domain of the CARs affected CAR Treg surface phenotype and functions such as cytokine production. Furthermore, signaling from the CD28 costimulation domain maintained CAR Treg suppressor function, whereas 4-1B costimulation did not. In vivo, CAR Tregs accumulated at sites expressing target antigen, and suppressed antigen specific effector T cell responses; however, only CAR Tregs with CD28 signaling domains were potent inhibitors of effector T cell mediated graft rejection in vivo. Our findings support the use of CD28 based CAR-Tregs for tissue specific immune suppression in the clinic.
Angela C. Boroughs, Rebecca C. Larson, Bryan D. Choi, Amanda A. Bouffard, Lauren S. Riley, Erik Schiferle, Anupriya S. Kulkarni, Curtis L. Cetrulo, David Ting, Bruce R. Blazar, Shadmehr Demehri, Marcela V. Maus
Costimulatory interactions control T cell activation at sites of activated antigen-presenting cells, including B cells. Blockade of the CD28/CD80/CD86 costimulatory axis with CTLA4Ig (abatacept) is widely used to treat certain autoimmune diseases. While transiently effective in subjects with new-onset type 1 diabetes (T1D), abatacept did not induce long-lasting immune tolerance. To elucidate mechanisms limiting immune tolerance in T1D, we performed unbiased analysis of whole blood transcriptomes and targeted measurements of cell subset levels in subjects from a clinical trial of abatacept in new-onset T1D. We showed that individual subjects displayed age-related immune phenotypes (“immunotypes”) at baseline, characterized by elevated levels of B cells or neutrophils, that accompanied rapid or slow progression, respectively, in both abatacept- and placebo-treated groups. A more pronounced immunotype was exhibited by a subset of subjects showing poor response (resistance) to abatacept. This resistance immunotype was characterized by a transient increase in activated B cells (one of the cell types that binds abatacept), reprogrammed costimulatory ligand gene expression, and reduced inhibition of anti-insulin antibodies. Our findings identify immunotypes in T1D subjects that are linked to the rate of disease progression, both in placebo- and abatacept-treated subjects. Furthermore, our results suggest therapeutic approaches to restore immune tolerance in T1D.
Peter S. Linsley, Carla J. Greenbaum, Cate Speake, S. Alice Long, Matthew J. Dufort
The rate of decline in insulin secretion after diagnosis with type 1 diabetes (T1D) varies substantially among individuals and with age at diagnosis, but the mechanism(s) behind this heterogeneity are not well understood. We investigated the loss of pancreatic β cell function in new-onset T1D subjects using unbiased whole blood RNA-seq and verified key findings by targeted cell count measurements. We found that patients who lost insulin secretion more rapidly had immune phenotypes (“immunotypes”) characterized by higher levels of B cells and lower levels of neutrophils, especially neutrophils expressing primary granule genes. The B cell and neutrophil immunotypes showed strong age dependence, with B cell levels in particular predicting rate of progression in young subjects only. This age relationship suggested that therapy targeting B cells in T1D would be most effective in young subjects with high pretreatment B cell levels, a prediction which was supported by data from a clinical trial of rituximab in new-onset subjects. These findings demonstrate a link between age-related immunotypes and disease outcome in new-onset T1D. Furthermore, our data suggest that greater success could be achieved by targeted use of immunomodulatory therapy in specific T1D populations defined by age and immune characteristics.
Matthew J. Dufort, Carla J. Greenbaum, Cate Speake, Peter S. Linsley
The auto antigen (Ag)-specific regulatory T cells (Tregs) from pluripotent stem cells (PSCs), i.e., PSC-Tregs, have the ability to suppress autoimmunity. PSC-Tregs can be programmed to be tissue-associated and to infiltrate into local inflamed tissues to suppress autoimmune responses after adoptive transfer. Nevertheless, the mechanisms by which the auto Ag-specific PSC-Tregs suppress the autoimmune response remain to be fully elucidated. In this study, we generated the functional auto Ag-specific Tregs from the induced PSC (iPSCs), i.e., iPSC-Tregs, and investigated the underlying mechanisms of autoimmunity suppression by these Tregs in a type 1 diabetes (T1D) murine model. A double transgenic (Tg) mouse model of T1D was established in F1 mice in which the first generation of RIP-mOVA Tg mice that were crossed with OT-I T cell receptor (TCR) Tg mice was challenged with vaccinia viruses expressing OVA (VACV-OVA). We show that adoptive transfer of OVA-specific iPSC-Tregs greatly suppressed autoimmunity in the animal model and prevented the insulin-secreting pancreatic β cells from destruction. Further, we demonstrate that the adoptive transfer significantly reduced the expression of ICAM-1 in the diabetic pancreas and inhibited the migration of pathogenic CD8+ T cells and the production of the pro-inflammatory IFN-γ in the pancreas. These results indicate that the stem cell-derived tissue-associated Tregs can robustly accumulate in the diabetic pancreas, and through down-regulating the expression of ICAM-1 in the local inflamed tissues and inhibiting the production of pro-inflammatory cytokine IFN-γ, suppress the migration and activity of the pathogenic immune cells that cause T1D.
Mohammad Haque, Fengyang Lei, Xiaofang Xiong, Jugal Kishore Das, Xingcong Ren, Deyu Fang, Shahram Salek-Ardakani, Jin-Ming Yang, Jianxun Song
Psoriasis (PS) is a systemic, immune-mediated inflammatory disorder. However, the whole lymphocyte compartment and the potential pathologies of PS have not been fully characterized. In the present study, we examined whole lymphocyte subsets and signal transduction proteins using high-dimensional single-cell mass cytometry and a bioinformatics pipeline for an in-depth characterization of the immune cell subsets and protein profiles involved in pathways in the peripheral blood of patients with PS. We identified 15 major immune cell populations in T cell lineages, and characterized various CD3+CD4+T helper and CD3+CD8+T cytotoxic cell populations simultaneously across 24 leukocyte markers and 7 proteins related to the signal transduction pathways. High-dimensional analysis identified three new subsets that are abundant in PS peripheral blood, resembling CD3-CD4+ lymphoid tissue inducer cells, Tc17, and CD8+CXCR3+ Tregs. We confirmed the CD3-CD4+ cells, and their features and functions, in an independent PS cohort. The use of single-cell mass cytometry allows systemic-level characterization of lymphocyte subpopulations and dysregulated signaling pathways in the blood of patients with PS, identifying abnormalities of different immune cell subsets. We validated that the CD3-CD4+ cells had elevated OX40 and decreased FRA2 expression, which were positively associated with the psoriasis area and severity index.
Ruru Guo, Ting Zhang, Xinyu Meng, Zhen Lin, Jinran Lin, Yu Gong, Xuesong Liu, Yuetian Yu, Guilin Zhao, Xianting Ding, Xiaoxiang Chen, Liangjing Lu
Th1 Tregs are characterized by the acquisition of proinflammatory cytokine secretion and reduced suppressor activity. Th1 Tregs are found at increased frequency in autoimmune diseases, including type 1 diabetes and multiple sclerosis (MS). We have previously reported that in vitro stimulation with IL-12 recapitulates the functional and molecular features of MS-associated Th1 Tregs, revealing a central role for hyperactivation of the Akt pathway in their induction. TIGIT is a newly identified coinhibitory receptor that marks Tregs that specifically control Th1 and Th17 responses. Here, we report that signaling through TIGIT counteracts the action of IL-12 in inducing the Th1 program. Specifically, TIGIT signaling represses production of IFN-γ and T-bet expression and restores suppressor function in Tregs treated with IL-12. FoxO1 functional inhibition abolishes the protective effect of TIGIT, indicating that TIGIT signaling promotes FoxO1 nuclear localization. Consistent with this observation, signaling through TIGIT leads to a rapid suppression of Akt function and FoxO1 phosphorylation. Finally, TIGIT stimulation reduces the production of IFN-γ and corrects the suppressor defect of Tregs from patients with MS. Our results indicate an important role for TIGIT in controlling the functional stability of Tregs through repression of Akt, suggesting that the TIGIT pathway could be targeted for immunomodulatory therapies in human autoimmune disorders.
Liliana E. Lucca, Pierre-Paul Axisa, Emily R. Singer, Neal M. Nolan, Margarita Dominguez-Villar, David A. Hafler
It has been reported that 2.5%–30% of human peripheral CD27– B cells are autoreactive and anergic based on unresponsiveness to antigen receptor (BCR) stimulation and autoreactivity of cloned and expressed BCR. The molecular mechanisms that maintain this unresponsiveness are unknown. Here, we showed that in humans anergy is maintained by elevated expression of PTEN, a phosphatidylinositol 3,4,5P-3-phosphatase. Upregulation of PTEN was associated with reduced expression of microRNAs that control its expression. Pharmacologic inhibition of PTEN lead to significant restoration of responsiveness. Consistent with a role in conferring risk of autoimmunity, B cells from type 1 diabetics and autoimmune thyroid disease patients expressed reduced PTEN. Unexpectedly, in healthy individuals PTEN expression was elevated in on average 40% of CD27– B cells, with levels gradually decreasing as IgM levels increase. Our findings suggest that a much higher proportion of the peripheral repertoire is autoreactive than previously thought and that B cells upregulate PTEN in a manner that is proportional to the recognition of autoantigens of increasing avidity, thus tuning BCR signaling to prevent development of autoimmunity while providing a reservoir of cells that can be readily activated to respond when needed.
Mia J. Smith, B. Rhodes Ford, Marynette Rihanek, Brianne M. Coleman, Andrew Getahun, Virginia D. Sarapura, Peter A. Gottlieb, John C. Cambier
Here, we report a pathogenic role for type I IFN (IFN-I) signaling in macrophages, and not β cells in the islets, for the development of type 1 diabetes (T1D). Following lymphocytic choriomeningitis (LCMV) infection in the Rip-LCMV-GP T1D model, macrophages accumulated near islets and in close contact to islet-infiltrating GP-specific (autoimmune) CD8+ T cells. Depletion of macrophages with clodronate liposomes or genetic ablation of Ifnar in macrophages aborted T1D, despite proliferation of GP-specific (autoimmune) CD8+ T cells. Histopathologically, disrupted IFNα/β receptor (IFNAR) signaling in macrophages resulted in restriction of CD8+ T cells entering into the islets with significant lymphoid accumulation around the islet. Collectively, these results provide evidence that macrophages via IFN-I signaling, while not entering the islets, are directly involved in interacting, directing, or restricting trafficking of autoreactive-specific T cells into the islets as an important component in causing T1D.
Brett S. Marro, Sarah Legrain, Brian C. Ware, Michael B.A. Oldstone
The antiinflammatory effects of i.v. Ig (IVIG) in the treatment of autoimmune disease are due, in part, to the Fc fragments of Ig aggregates. In order to capitalize on the known antiinflammatory and tolerogenic properties of Ig Fc aggregates, we created a recombinant human IgG1 Fc multimer, GL-2045. In vitro, GL-2045 demonstrated high-avidity binding to Fc receptors, blocked the binding of circulating immune complexes from patients with rheumatoid arthritis to human Fcγ receptors (FcγRs), and inhibited antibody-mediated phagocytosis at log order–lower concentrations than IVIG. In vivo, administration of GL-2045 conferred partial protection against antibody-mediated platelet loss in a murine immune thrombocytopenic purpura (ITP) model. GL-2045 also suppressed disease activity in a therapeutic model of murine collagen-induced arthritis (CIA), which was associated with reduced circulating levels of IL-6. Furthermore, GL-2045 administration to nonhuman primates (NHPs) transiently increased systemic levels of the antiinflammatory cytokines IL-10 and IL-1RA, reduced the proinflammatory cytokine IL-8, and decreased surface expression of CD14 and HLA-DR on monocytes. These findings demonstrate the immunomodulatory properties of GL-2045 and suggest that it has potential as a treatment for autoimmune and inflammatory diseases, as a recombinant alternative to IVIG.
Xiaoyu Zhang, Jane Owens, Henrik S. Olsen, Edward So, Erin Burch, Mark C. McCroskey, Xianfeng Li, Gregory L. Weber, Donald Bennett, Denis Rybin, Hua Zhou, Haiping Hao, Emmanuel Y. Mérigeon, David S. Block, Gregory LaRosa, Scott E. Strome
Systemic lupus erythematosus (SLE) is a highly variable autoimmune disease that can involve severe organ-threatening symptoms, such as lupus nephritis. Certain drugs, such as mycophenolate mofetil (MMF), are effective at reducing morbidity associated with nephritis; however, the immune pathways associated with disease suppression are poorly defined. Here, we provide evidence that MMF inhibits phosphorylation of STAT3 and other associated immune pathways. Using mass cytometry and bead-based or ELISA assays, the systemic phenotype of SLE patients not taking (MMF–) or taking (MMF+) MMF were studied. MMF+ SLE patients had significant reductions in total numbers of transitional B cells, plasmablasts, and T cells, specifically CD4+ Th17-type and CD4+ Treg-type cells, compared with MMF– patients. Plasma soluble mediators were decreased in MMF+ patients including chemokines (MIG/CXCL9 and SDF-1α/CXCL12) and growth factors (VEGF-A and PDGF-BB). Soluble mediators and cell subsets grouped by functional properties revealed significant modifications associated with STAT3 and B cell pathways. Further, healthy PBMCs treated with IL-6 revealed a reduction in p-STAT3 following the addition of mycophenolic acid (the active metabolite of MMF). In conclusion, the inhibition of STAT3 phosphorylation by MMF may explain the effectiveness of this treatment in SLE patients, since increased levels of p-STAT3 are associated with disease pathology.
Samantha Slight-Webb, Joel M. Guthridge, Eliza F. Chakravarty, Hua Chen, Rufei Lu, Susan Macwana, Krista Bean, Holden T. Maecker, Paul J. Utz, Judith A. James
No posts were found with this tag.